The use of NEG pumps and coatings in large vacuum systems: experience and limitations

Today Non Evaporable Getter (NEG) materials are commonly used for vacuum production in large vacuum systems, in particular in particle accelerators. Recently, a new technology based on NEG sputtering techniques was successfully developed by CERN and utilized at various synchrotron light sources and ion collider facilities. The aim was to improve the vacuum performance in some conductance limited vacuum systems in order to minimize the gas bremsstrahlung emission and to reduce the impact of the electron-cloud instability discovered in some machines. The most recent results obtained are presented

Antimicrobial susceptibility to zinc bacitracin of Clostridium perfringens of rabbit origin

Zinc bacitracin is widely used in Italian rabbit farms to control both Epizootic Rabbit Enteropathy (ERE) and clostridiosis, and field results demonstrate useful activity. Nevertheless, data regarding the in vitro efficacy of zinc bacitracin against clostridia of rabbit origin are not available. In this study, the minimal inhibitory concentrations (MICs) of zinc bacitracin were evaluated in 123 C. perfringens strains isolated from rabbits in Italian fattening units. The agar dilution method was performed in Brucella Agar supplemented with laked sheep blood, haemin and vitamin K1, as recommended in NCCLS document M11-A6. Most strains (94.3%) had low MIC values (£ 0.5 mg/ml), and a few strains (4%) were inhibited by a concentration of 1 mg/ml. Two isolates (1.6%) had a MIC value of 16mg/ml. The MIC values of ATCC reference strains showed a good fit between each batch. MIC required to inhibit the 90% of organisms was 0.5 mg/ml and the presence of only two strains with MIC=16 mg/ml revealed the susceptibility to zinc bacitracin of Italian isolates of C. perfringens from rabbit and the absence of acquired resistance.Agnoletti, F.; Bacchin, C.; Bano, L.; Passera, A.; Favretti, M.; Mazzolini, E. (2007). Antimicrobial susceptibility to zinc bacitracin of Clostridium perfringens of rabbit origin. World Rabbit Science. 15(1):19-22. doi:10.4995/wrs.2007.609192215

The liver, liver metastasis and liver cancer: A special case for immunotherapy with cytokines and immunostimulatory monoclonal antibodies

"The immunologic outcome of liver transplantations tells us that this organ is somehow specialized as a tolerogenic organ under steady state conditions."Fil: Mazzolini Rizzo, Guillermo Daniel. Universidad Austral. Facultad de Ciencias Biomédicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Ochoa, María C. Universidad de Navarra; EspañaFil: Morales Kastresana, Aizea. Universidad de Navarra; EspañaFil: Sanmamed, Miguel F.. Universidad de Navarra; EspañaFil: Melero, Ignacio. Universidad de Navarra; Españ

Conformational rearrangements in the transmembrane domain of CNGA1 channels revealed by single-molecule force spectroscopy

Cyclic nucleotide-gated (CNG) channels are activated by binding of cyclic nucleotides. Although structural studies have identified the channel pore and selectivity filter, conformation changes associated with gating remain poorly understood. Here we combine single-molecule force spectroscopy (SMFS) with mutagenesis, bioinformatics and electrophysiology to study conformational changes associated with gating. By expressing functional channels with SMFS fingerprints in Xenopus laevis oocytes, we were able to investigate gating of CNGA1 in a physiological-like membrane. Force spectra determined that the S4 transmembrane domain is mechanically coupled to S5 in the closed state, but S3 in the open state. We also show there are multiple pathways for the unfolding of the transmembrane domains, probably caused by a different degree of \u3b1-helix folding. This approach demonstrates that CNG transmembrane domains have dynamic structure and establishes SMFS as a tool for probing conformational change in ion channels

Acceleration of TAA-induced liver fibrosis by stress exposure is associated with upregulation of nerve growth factor and glycopattern deviations

Liver fibrosis results from many chronic injuries and may often progress to cirrhosis and hepatocellular carcinoma (HCC). In fact, up to 90% of HCC arise in a cirrhotic liver. Conversely, stress is implicated in liver damage, worsening disease outcome. Hence, stress could play a role in disrupting liver homeostasis, a concept that has not been fully explored. Here, in a murine model of TAA-induced liver fibrosis we identified nerve growth factor (NGF) to be a crucial regulator of the stress-induced fibrogenesis signaling pathway as it activates its receptor p75 neurotrophin receptor (p75NTR), increasing liver damage. Additionally, blocking the NGF decreased liver fibrosis whereas treatment with recombinant NGF accelerated the fibrotic process to a similar extent than stress challenge. We further show that the fibrogenesis induced by stress is characterized by specific changes in the hepatoglycocode (increased β1,6GlcNAc-branched complex N-glycans and decreased core 1 O-glycans expression) which are also observed in patients with advanced fibrosis compared to patients with a low level of fibrosis. Our study facilitates an understanding of stress-induced liver injury and identify NGF signaling pathway in early stages of the disease, which contributes to the established fibrogenesis

Intratumoral injection of dendritic cells engineered to secrete interleukin-12 by recombinant adenovirus in patients with metastatic gastrointestinal carcinomas.

PURPOSE: To evaluate the feasibility and safety of intratumoral injection of autologous dendritic cells (DCs) transfected with an adenovirus encoding interleukin-12 genes (AFIL-12) for patients with metastatic gastrointestinal carcinomas. Secondarily, we have evaluated biologic effects and antitumoral activity. PATIENTS AND METHODS: Seventeen patients with metastatic pancreatic (n = 3), colorectal (n = 5), or primary liver (n = 9) malignancies entered the study. DCs were generated from CD14+ monocytes from leukapheresis, cultured and transfected with AFIL-12 before administration. Doses from 10 x 10(6) to 50 x 10(6) cells were escalated in three cohorts of patients. Patients received up to three doses at 21-day intervals. RESULTS: Fifteen (88%) and 11 of 17 (65%) patients were assessable for toxicity and response, respectively. Intratumoral DC injections were mainly guided by ultrasound. Treatment was well tolerated. The most common side effects were lymphopenia, fever, and malaise. Interferon gamma and interleukin-6 serum concentrations were increased in 15 patients after each treatment, as well as peripheral blood natural killer activity in five patients. DC transfected with AFIL-12 stimulated a potent antibody response against adenoviral capsides. DC treatment induced a marked increase of infiltrating CD8+ T lymphocytes in three of 11 tumor biopsies analyzed. A partial response was observed in one patient with pancreatic carcinoma. Stable disease was observed in two patients and progression in eight patients, with two of the cases fast-progressing during treatment. CONCLUSION: Intratumoral injection of DC transfected with an adenovirus encoding interleukin-12 to patients with metastatic gastrointestinal malignancies is feasible and well tolerated. Further studies are necessary to define and increase clinical efficacy

Gene expression profiling reveals a conserved microglia signature in larval zebrafish

Microglia are the resident macrophages of the brain. Over the past decade, our understanding of the function of these cells has significantly improved. Microglia do not only play important roles in the healthy brain but are involved in almost every brain pathology. Gene expression profiling allowed to distinguish microglia from other macrophages and revealed that the full microglia signature can only be observed in vivo. Thus, animal models are irreplaceable to understand the function of these cells. One of the popular models to study microglia is the zebrafish larva. Due to their optical transparency and genetic accessibility, zebrafish larvae have been employed to understand a variety of microglia functions in the living brain. Here, we performed RNA sequencing of larval zebrafish microglia at different developmental time points: 3, 5, and 7 days post fertilization (dpf). Our analysis reveals that larval zebrafish microglia rapidly acquire the core microglia signature and many typical microglia genes are expressed from 3 dpf onwards. The majority of changes in gene expression happened between 3 and 5 dpf, suggesting that differentiation mainly takes place during these days. Furthermore, we compared the larval microglia transcriptome to published data sets of adult zebrafish microglia, mouse microglia, and human microglia. Larval microglia shared a significant number of expressed genes with their adul

SPARC (secreted protein acidic and rich in cysteine) knockdown protects mice from acute liver injury by reducing vascular endothelial cell damage

Secreted protein, acidic and rich in cysteine (SPARC) is involved in many biological process including liver fibrogenesis, but its role in acute liver damage is unknown. To examine the role of SPARC in acute liver injury, we used SPARC knock-out (SPARC−/−) mice. Two models of acute liver damage were used: concanavalin A (Con A) and the agonistic anti-CD95 antibody Jo2. SPARC expression levels were analyzed in liver samples from patients with acute-on-chronic alcoholic hepatitis (AH). SPARC expression is increased on acute-on-chronic AH patients. Knockdown of SPARC decreased hepatic damage in the two models of liver injury. SPARC−/− mice showed a marked reduction in Con A-induced necroinflammation. Infiltration by CD4+ T cells, expression of tumor necrosis factor-α and interleukin-6 and apoptosis were attenuated in SPARC−/− mice. Sinusoidal endothelial cell monolayer was preserved and was less activated in Con A-treated SPARC−/− mice. SPARC knockdown reduced Con A-induced autophagy of cultured human microvascular endothelial cells (HMEC-1). Hepatic transcriptome analysis revealed several gene networks that may have a role in the attenuated liver damaged found in Con A-treated SPARC−/− mice. SPARC has a significant role in the development of Con A-induced severe liver injury. These results suggest that SPARC could represent a therapeutic target in acute liver injury

Dendritic Cells Take up and Present Antigens from Viable and Apoptotic Polymorphonuclear Leukocytes

Dendritic cells (DC) are endowed with the ability to cross-present antigens from other cell types to cognate T cells. DC are poised to meet polymorphonuclear leukocytes (PMNs) as a result of being co-attracted by interleukin-8 (IL-8), for instance as produced by tumor cells or infected tissue. Human monocyte-derived and mouse bone marrow-derived DC can readily internalize viable or UV-irradiated PMNs. Such internalization was abrogated at 4°C and partly inhibited by anti-CD18 mAb. In mice, DC which had internalized PMNs containing electroporated ovalbumin (OVA) protein, were able to cross-present the antigen to CD8 (OT-1) and CD4 (OT-2) TCR-transgenic T cells. Moreover, in humans, tumor cell debris is internalized by PMNs and the tumor-cell material can be subsequently taken up from the immunomagnetically re-isolated PMNs by DC. Importantly, if human neutrophils had endocytosed bacteria, they were able to trigger the maturation program of the DC. Moreover, when mouse PMNs with E. coli in their interior are co-injected in the foot pad with DC, many DC loaded with fluorescent material from the PMNs reach draining lymph nodes. Using CT26 (H-2d) mouse tumor cells, it was observed that if tumor cells are intracellularly loaded with OVA protein and UV-irradiated, they become phagocytic prey of H-2d PMNs. If such PMNs, that cannot present antigens to OT-1 T cells, are immunomagnetically re-isolated and phagocytosed by H-2b DC, such DC productively cross-present OVA antigen determinants to OT-1 T cells. Cross-presentation to adoptively transferred OT-1 lymphocytes at draining lymph nodes also take place when OVA-loaded PMNs (H-2d) are coinjected in the footpad of mice with autologous DC (H-2b). In summary, our results indicate that antigens phagocytosed by short-lived PMNs can be in turn internalized and productively cross-presented by DC